Time series analysis demonstrates the absence of pulsatile hyphal growth.
Hyphal tip growth has been previously reported as pulsatile, defined as regularly alternating fast and slow rates of extension. The growth of pollen tubes, and hyphae of Neurospora crassa and Saprolegnia ferax were analysed using high spatial and temporal resolution. By using long (100-500 s) records of growth rate, sampled every second, it was possible to apply rigorous statistical analysis of the time series. As previously demonstrated, pollen tubes can show pulsatile growth, detectable with this system. In contrast, hyphal growth rates do not show any evidence of pulsatile growth; instead, growth rates appear to fluctuate randomly. It is concluded that pulsatile growth is not a common feature of hyphal tip growth. (+info)
A novel omega3-fatty acid desaturase involved in the biosynthesis of eicosapentaenoic acid.
Long-chain n-3 PUFAs (polyunsaturated fatty acids) such as EPA (eicosapentaenoic acid; 20:5 n-3) have important therapeutic and nutritional benefits in humans. In plants, cyanobacteria and nematodes, omega3-desaturases catalyse the formation of these n-3 fatty acids from n-6 fatty acid precursors. Here we describe the isolation and characterization of a gene ( sdd17 ) derived from an EPA-rich fungus, Saprolegnia diclina, that encodes a novel omega3-desaturase. This gene was isolated by PCR amplification of an S. diclina cDNA library using oligonucleotide primers corresponding to conserved regions of known omega3-desaturases. Expression of this gene in Saccharomyces cerevisiae, in the presence of various fatty acid substrates, revealed that the recombinant protein could exclusively desaturate 20-carbon n-6 fatty acid substrates with a distinct preference for ARA (arachidonic acid; 20:4 n-6), converting it into EPA. This activity differs from that of the known omega3-desaturases from any organism. Plant and cyanobacterial omega3-desaturases exclusively desaturate 18-carbon n-6 PUFAs, and a Caenorhabditis elegans omega3-desaturase preferentially desaturated 18-carbon PUFAs over 20-carbon substrates, and could not convert ARA into EPA when expressed in yeast. The sdd17 -encoded desaturase was also functional in transgenic somatic soya bean embryos, resulting in the production of EPA from exogenously supplied ARA, thus demonstrating its potential for use in the production of EPA in transgenic oilseed crops. (+info)
Expressed sequence tags from the oomycete fish pathogen Saprolegnia parasitica reveal putative virulence factors.
BACKGROUND: The oomycete Saprolegnia parasitica is one of the most economically important fish pathogens. There is a dramatic recrudescence of Saprolegnia infections in aquaculture since the use of the toxic organic dye malachite green was banned in 2002. Little is known about the molecular mechanisms underlying pathogenicity in S. parasitica and other animal pathogenic oomycetes. In this study we used a genomics approach to gain a first insight into the transcriptome of S. parasitica. RESULTS: We generated 1510 expressed sequence tags (ESTs) from a mycelial cDNA library of S. parasitica. A total of 1279 consensus sequences corresponding to 525944 base pairs were assembled. About half of the unigenes showed similarities to known protein sequences or motifs. The S. parasitica sequences tended to be relatively divergent from Phytophthora sequences. Based on the sequence alignments of 18 conserved proteins, the average amino acid identity between S. parasitica and three Phytophthora species was 77% compared to 93% within Phytophthora. Several S. parasitica cDNAs, such as those with similarity to fungal type I cellulose binding domain proteins, PAN/Apple module proteins, glycosyl hydrolases, proteases, as well as serine and cysteine protease inhibitors, were predicted to encode secreted proteins that could function in virulence. Some of these cDNAs were more similar to fungal proteins than to other eukaryotic proteins confirming that oomycetes and fungi share some virulence components despite their evolutionary distance CONCLUSION: We provide a first glimpse into the gene content of S. parasitica, a reemerging oomycete fish pathogen. These resources will greatly accelerate research on this important pathogen. The data is available online through the Oomycete Genomics Database. (+info)
Effects of nitrate and the pathogenic water mold Saprolegnia on survival of amphibian larvae.
We tested for a synergism between nitrate and Saprolegnia, a pathogenic water mold, using larvae of 3 amphibian species: Ambystoma gracile (northwestern salamander), Hyla regilla (Pacific treefrog) and Rana aurora (red-legged frog). Each species was tested separately, using a 3 x 2 fully factorial experiment with 3 nitrate treatments (none, low and high) and 2 Saprolegnia treatments (Saprolegnia and control). Survival of H. regilla was not affected significantly by either experimental factor. In contrast, survival of R. aurora was affected by a less-than-additive interaction between Saprolegnia and nitrate. Survival of R. aurora was significantly lower in the Saprolegnia compared to the control treatment when nitrate was not added, but there was no significant difference in survival between Saprolegnia and control treatments in the low and high nitrate treatments, consistent with increased nitrate preventing Saprolegnia from causing mortality of R. aurora. Survival of A. gracile followed a similar pattern, but the difference between Saprolegnia and control treatments when nitrate was not added was not significant, nor was the nitrate x Saprolegnia interaction. Our study suggests that Saprolegnia can cause mortality in amphibian larvae, that there are interspecific differences in susceptibility and that the effects of Saprolegnia on amphibians are context-dependent. (+info)
Antibody response of brown trout Salmo trutta injected with pathogenic Saprolegnia parasitica antigenic extracts.
Brown trout Salmo trutta injected with antigenic extracts from a pathogenic isolate of Saprolegnia parasitica developed specific antibodies that were detected by enzyme-linked immunosorbent assay (ELISA), immunofluorescence (IF) and Western blotting (WB), but not by immunodiffusion (ID). Three groups of five 2 yr old brown trout were injected intraperitoneally with 3 different antigenic extracts: small hyphal fragments (HF) and soluble extracts from sonicated mycelia grown in medium with or without beta-sytosterol (SEB and SE, respectively). In the 2 groups injected with SE and SEB, antibodies were found in 66.7 % of the serum samples by ELISA, 54.5% by IF and 48.5% by WB. In the group injected with HF, only 1 trout survived the experiment, and in this fish only 1 sample was positive by ELISA. The results obtained by ELISA and IF were similar and show that there is cross-reaction between the antigens used. By WB, the proteins most frequently recognised were 2 proteins of 25 and 29 kDa. No significant differences were found in the groups injected with SE or SEB. (+info)
Species boundaries within Saprolegnia (Saprolegniales, Oomycota) based on morphological and DNA sequence data.
Saprolegnia is a common and widespread genus of Oomycetes, however species identifications are difficult and uncertain. To test whether keys based on morphological characters could identify species as determined by molecular characters we determined partial DNA sequences for the 28S rRNA gene and the complete internal transcribed spacer (ITS) region for 55 isolates belonging to Saprolegnia and one isolate of Protoachlya hypogyna that exhibited saprolegnoid zoospore discharge in water culture. Phylogenetic analyses of the combined sequence data yielded 10 robustly supported clades that probably represent separate species. Morphological analyses of all isolates revealed that each DNA-based clade could be delimited from others by autapomorphic or unique combinations of morphological character states but not without employing several features previously not used at the species level. Taxonomic implications of these results are discussed and recommendations for less equivocal characterization of new Saprolegnia species are made. (+info)
A genetic component of resistance to fungal infection in frog embryos.
Saprolegniaceae identified on amphibian eggs throughout the Pacific Northwest, USA, by internal transcribed spacer sequences and phylogenetic analysis.
We assessed the diversity and phylogeny of Saprolegniaceae on amphibian eggs from the Pacific Northwest, with particular focus on Saprolegnia ferax, a species implicated in high egg mortality. We identified isolates from eggs of six amphibians with the internal transcribed spacer (ITS) and 5.8S gene regions and BLAST of the GenBank database. We identified 68 sequences as Saprolegniaceae and 43 sequences as true fungi from at least nine genera. Our phylogenetic analysis of the Saprolegniaceae included isolates within the genera Saprolegnia, Achlya and Leptolegnia. Our phylogeny grouped S. semihypogyna with Achlya rather than with the Saprolegnia reference sequences. We found only one isolate that grouped closely with S. ferax, and this came from a hatchery-raised salmon (Idaho) that we sampled opportunistically. We had representatives of 7-12 species and three genera of Saprolegniaceae on our amphibian eggs. Further work on the ecological roles of different species of Saprolegniaceae is needed to clarify their potential importance in amphibian egg mortality and potential links to population declines. (+info)